Liver microsomal steroid hydroxylation activities, which are catalyzed by specific forms of cytochrome P-450s, are known to exhibit sex-dependent expression. In inbred mice, the expression of steroid hydroxylations differs from one to another strain. It has been known that 16Alpha-hydroxylation activity of female mice is recessively regulated by a single gene between 129/J and C56BL/6J. This genetic regulation, however, between 129/J and BACB/CJ was found to be autosomal dominant, indicating that more than one allele are involved in this regulation. The sex-dependent expression of hydroxylases in adulthood are irreversibly predetermined by neonatal androgens. It is reasonable to assume that these differences of microsomal cytochrome P-450s in sex may effect sex-dependent drug toxicity, including chemical carcinogenesis. To understand the biological phenomena surrounding sex-dependent and developmental regulation of gene expression in steroid hydroxylases, we have previously purified female predominant mouse testosterone 15a-hydroxylase P450(15a). Now male predominant mouse testosterone 16a-hydrocylase has been purified and characterized. A specific antibody against P450(15a) or P450-16a was raised in rabbits, and they completely inhibited only 16a - or 15a-hydroxylation activity. Liver polysomes bearing P-450(15a) or P-450(16a) mRNA were precipitated by the antibodies 32P- from which the cDNA banks were constructed. By the selective hybridization to labeled probe synthesized from the immunoenriched mRNAs, the cDNAs encoding P-450(16a) has been isolated. By the further characterization and isolation of genomic clones with these cDNA probes, we hope to understand the mechanisms of sex- and age-dependent gene expression of these hydroxylases, and gain insight into the role of cytochrome P-450s in steroid metabolisms and sex differences of chemical toxicity. We have also isolated and characterized cDNAs for drug-induced mouse liver microsomal cytochrome P-450. A comparison between androgen-dependent and xenobiotic-dependent cytochrome P-450 genes will provide useful and better information to understand mechanisms of gene expression.